Sizova, Irina and Fuhrmann, Markus and Hegemann, Peter (2001) A Streptomyces rimosus aphVIII gene coding for a new type phosphotransferase provides stable antibiotic resistance to Chlamydomonas reinhardtii. GENE, 277 (1-2). pp. 221-229. ISSN 0378-1119
Full text not available from this repository. (Request a copy)Abstract
Although Chlamydomonas reinhardtii serves as the most popular algal model system, no efficient enzymatic selection marker for the nuclear transformation of wild-type cells is available. We sequenced an aminoglycoside 3'-phosphotransferase gene (aph) from Streptomyces rimosus. Though the derived protein sequence is homologous to members of APH type V, it constitutes a new type, named APHVIII Since the aphVIII gene has a codon bias similar to that of the nuclear genome of green algae, the aphVIII coding sequence was fused to the 5'-and 3'-untranslated regions of the C reinhardtii rbcS2 gene. C. reinhardtii transformants were capable of inactivating the antibiotics paromomycin, kanamycin, and neomycin, to which wild-type cells are sensitive. After addition of the 5'-region of hsp70A as a second promoter and insertion of the rbcS2 intron I, the transformation rate increased to two transformants per I X 10(5) cells, which is close to the efficiency of transforming auxotrophic strains with the homologous marker arg7. Transformation with the promoter-less aphVIII led to random gene fusion at high frequency. In an aphVIII-based reporter gene assay we have found a so far unknown promoter activity of the 3'-untranslated region of rbcS2, that may promote antisense RNA synthesis from the rbcS2 gene in vivo. We conclude that the aphVIII gene is a useful marker for nuclear transformation and promoter tagging of C. reinhardtii wild-type and probably other green algae. (C) 2001 Published by Elsevier Science B.V.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | DOMINANT SELECTABLE MARKER; NUCLEAR TRANSFORMATION; FOREIGN GENE; EXPRESSION; PROTEINS; VECTORS; FUSION; TOOL; DNA; antisense; green algae; paromomycin; rbcS2 |
| Subjects: | 500 Science > 570 Life sciences |
| Divisions: | Biology, Preclinical Medicine > Institut für Biochemie, Genetik und Mikrobiologie > Lehrstuhl für Biochemie I |
| Depositing User: | Dr. Gernot Deinzer |
| Date Deposited: | 01 Dec 2021 15:38 |
| Last Modified: | 01 Dec 2021 15:38 |
| URI: | https://pred.uni-regensburg.de/id/eprint/41030 |
Actions (login required)
 |
View Item |
